Neisseria gonorrhoeae is a microbe of many paradoxes. The difference in presentation of infection in men and women is one of the most obvious. Ninety-five percent of infections in male are symptomatic four to thirty days after exposure. By contrast, the majority of cases (40-60%) of cervical infection are asymptomatic. Many of these women do not receive treatment and serve as silent reservoirs of infection. In a high percentage of infected women, asymptomatic cervical gonorrhea will eventually become symptomatic and involve the upper genital tract with consequences for PID, chronic pelvic pain and, in some, sterility. Females with asymptomatic gonorrhea are the persistent source of infection in any community. It has been proposed that the association of gonococci with cervical squamous cells may provide a protected environment for the continuation of infection. Recently, we have made the observation that during human infection, expression of the Gal-CicNAc LOS epitope is increased ten-fold when compared to the expression observed in plate grown organisms. This epitope is heavily sialylated during human infection and could protect the organism by what has been described as a sialylated "biologic mask". This concept is supported by an increasing number of studies that suggest that sialylated LOS enhances the ability of the gonococcus to evade human bactericidal antibody and oposonphagocytosis. We have established a model system in which Hec1B cells have been chronically infected (for up to eight weeks) with Neisseria gonorrhoeae. Studies of the LOS expressed by organisms in this model system indicate that the Ga1-G1cNAc epitope expression is increased to levels seen during natural infection. In addition, this epitope is sialylated. Thus, we now have a model to study changes in LOS expression that we have observed in infected exudates. We have recently cloned and defined two gonococcal LOS biosynthesis genes that encode for phosphoglucomutase (pgm) and UDP-galactose-4-epimerase (galE). We propose to continue to work on defining LOS oligosaccharide biosynthesis at the genetic level. In addition, we will study the hypothesis that gonococcal LOS expression is regulated differently during in vitro and in vivo growth. This will be accomplished by the following specific aims: 1) To clone and define gene clusters responsible for LOS biosynthesis, 2) To define the gonococcal LOS biosynthesis gene cluster associated with pgm and galE, and 3) To study the regulation of LOS biosynthesis at the genetic level in N. gonorrhoeae strain 1291 after in vitro culture, in the Hec1B cell line chronically infected with N. gonorrhoeae and in human exudates from patients with gonococcal urethritis and cervicitis.